Ion-pair high-pressure liquid chromatography of cis-trans isomers of retinoic acid in tissues of vitamin A-sufficient rats.

Abstract

Naturally occurring retinoids were separated by reversed-phase high-pressure liquid chromatography on an octadecylsilane column eluted with acetonitrile-potassium phosphate buffer (pH 7.2) mixtures. The order of elution from a mixture of 500 ng each of the following standards was 4-oxo retinoic acid (RA), retinyl phosphate (RP), 13-cis RA, all-trans RA, retinol, retinal, retinyl acetate, anhydroretinol, and retinyl palmitate. This method was employed to investigate the cis-trans isomerization of RA and its metabolism in vitamin A-sufficient male rats. Rats (200 g) were injected intraperitoneally with 50 muCi of either [10-3H]-all-trans RA (5.4 micrograms) or [11-3H]-13-cis RA (8.8 micrograms) and killed after 0.5 and 3 hr. Blood, liver, kidney, small intestines, and testes were removed and lyophilized. All-trans RA was converted at 0.5 hr after injection to 13-cis RA in all the tissues examined, with the exception of the small intestine; the conversion ranged from 2.4 to 6.9% of the total radioactivity. In addition, all-trans RA was converted to metabolites (17.5--47.7%) of greater polarity than 4-oxo RA. After 3 hr, most of the radioactivity (75--90%) was found in the highly polar metabolites. 13-cis RA was also partially isomerized to the all-trans RA and to the highly polar metabolites by 0.5 and 3 hr after injection. Appreciable radioactivity (10--41%) still resided in the 13-cis RA fraction after 3 hr. These results indicate that 13-cis RA is partially isomerized to all-trans RA and that all-trans RA is rapidly metabolized to highly polar compounds in tissues of vitamin A-sufficient rats.